Luteinizing hormone(LH) and human chorionic gonadotropin (hCG) bind to the same receptor (LHR). These hormones and LHR play crucial roles in reproduction, women's health and some ovarian and prostate cancers. LHR and hormone derivatives have been used or are tested for diagnosis and therapeutics. Our long term goal is to understand the molecular mechanism of LHR's interaction with hCG and generation of differential signals for second messengers, cAMP and inositol phosphates (IP). LHR and other glycoprotein hormone receptors belong to a structurally unique subfamily of G-protein-coupled receptors. Each receptor consist of the extracellular N-terminal half with approximately 350 amino acids (exodomain) and the membrane associated C-terminal half with an equal number of amino acids (endodomain). The exodomain is generally thought to form a 1/3 donut structure by 7-9 leu Rich Repeats (LRR) and to bind hCG via the LRRs. Yet, little experimental evidence is available to support the LRRs. Since we and others reported, nearly ten years ago, that the exodomain is capable of high affinity binding site for hCG, that the hCG/exodomain complex undergoes a conformational adjustment upon hormone binding, and that the endodomain is the site for signal generation. Based on these observations we proposed that the initial hormone/exodomain complex interacts with the endodomain and this secondary interaction is responsible for signal generation. Lately, many of these propositions have been demonstrated and adopted by others. During the current grant period, we have made a number of significant new observations, furthering the current understanding, models and hypotheses as well as introducing new concepts: The LRRs are active in LHR and LRR4 interacts hCG, the N and C-flanking regions of the LRRs appear to reach the back of hCG and wrap around it (which may be key factor in the high binding affinity), and the C-terminal region of the LRRs seems to activate the endodomain to generate signal, whereas LRR4 suppresses the endodomain. On the other hand, the 3 exoloops share common and distinct functions. The exoloop 3 is involved in differentially generating signals for cAMP and Ips, the exoloop 2 appears to be the primary site for contacting the exodomain, while the exoloop 1 may be involved in inter-exoloop interaction and signal generation. This is a mini-breakthrough. In addition, we have observed that the hCG/exodomain complex of one LHR may contact and activate the endodomain of another LHR, which has a far reaching implication on signal transduction in general. We propose to extend our observations (Specific Aim 1) to determine the interaction of the exodomain with hCG and (Specific Aim 2) to determine the interactions and function of the three exoloops. When successfully carried out, these studies will provide new insights on the hormone binding and intra- and inter- molecular, differential signal generation for LHR and other receptors, and therefore, clinical and industrial applications.